1. Field of the Invention
The present invention relates to a micro analytical method, a sampling plate used in the same, a method of detecting organic compounds by the use of the micro analytical method, and an apparatus for the same.
2. Description of the Prior Art
Micro analytical methods using, for example, a microscopic FTIR (Fourier Transform Infrared Ray Spectrophotometer), a FTIR with a beam condenser, a FTIR with a condensing lens and the like have been suitable for an analysis of a very small quantity of minute organic matter, but a disadvantage has occurred in the conventional means for condensing a very small quantity of sample in that a condensed sample is diffused to reduce the thickness thereof, thereby reducing the sensitivity.
That is to say, a very small quantity of solution of a solute in a solvent has been deposited drop by drop on a mirror-finished metallic base by the use of an instrument, such as a riffle sampler, and the solvent in the solution has been evaporated to condense the solute on the base, which condensed solution has been used as the sample.
However, by this condensing means, the solution deposited drop by drop on the base has been expanded all over said mirror-finished surface of the base and diffused in the form of islands to be condensed, so that the thickness of said sample has been apt to be reduced.
More concretely speaking, in the case where 1 .mu.l of a solution of 10 .mu.l of fluid paraffin in 100 ml of acetone has been deposited drop by drop on the base, the solution has been immediately expanded to a circle having a diameter of 3 to 5 mm. The solvent begins to evaporate from a circumferential portion of said circle and at the same time the solute is turned into numerous islands. These islands of solute are condensed in the form of a ring having a diameter of 3 to 5 mm, whereby the thickness of the sample is remarkably reduced.
In this case, the absorption of infrared rays was insufficient. Concretely speaking, a transmission factor of infrared rays measured by a microscopic FTIR/FT-530 (made by HORIBA, Ltd.) was about 98% at a wave number of about 1,450 cm.sup.-1 as shown in FIG. 38. That is to say, the absorbing degree of light is remarkably low to an extent of 0.01 and thus a highly accurate analysis by the microscopic spectrometric analytical method was difficult.
In addition, the above-described expansion of the solution has no orientation, so that it has been difficult to fix positions where the solute is to be condensed in an appointed manner. This has led to difficulty in the automation of the spectrometric analysis.
Furthermore, disadvantages have occurred in that if positions, where the solution is deposited drop by drop, are allowed to approach so that the solution may be deposited drop by drop at many points compared with the size of a sample table, the solutions deposited drop by drop are brought into contact with each other according to the expansion thereof or the solution deposited drop by drop is attracted to the already condensed sample so as to be brought into contact with it.
By the way, after FTIR has been widely used recently, the so-called HPLC/FTIR, in which a solvent in a solution eluted from HPLC (high-speed liquid chromatography) is removed to be used as a sample in FTIR, has been proposed to be practically used.
Methods of measuring as the above described HPLC/FTIR include for example
(1) A method in which an eluted solution is heated, concentrated and removed in sequential phase mode and then a solute is measured by the diffusion-reflection method or the transmission method;
(2) A method in which water is replaced by acetone and methanol by the use of 2,2-dimethoxypropane and acids in an inverted phase mode and acetone and methanol are heated to evaporatively remove solvents and then a solute is measured by the diffusion-reflection method or the transmission method;
(3) A method in which a solute is extracted by the use of chloroform or dichloromethane having a comparatively narrow absorption band in a sequential phase mode to measure a spectrum thereof by the transmission method in the form of liquid or a solvent is evaporatively removed and then a spectrum of a solute is measured by the diffusion-reflection method or the transmission method; and the like.
In any of the above described methods, the sample is irradiated with infrared rays and an absorption spectrum resulting from a vibration causing a change in a dipole moment of molecular vibrations is measured. In these methods, an infrared absorption is characterized in having absorption wavelength bands peculiar to organic functional groups and thus can be applied for qualitatively or quantitatively determining organic functional groups, discriminating similar compounds, making reaction mechanisms clear and analyzing structures, and additionally can analyze a multi-component system comprising at least several kinds of components. However, disadvantages have occurred in that not only is the sensitivity several hundred ng (10.sup.-9 g) and thus it is unsuitable for a measurement of a very small quantity of component, but also HPLC and the exclusive FTIR are required and thus an apparatus is large-scaled and expensive.
In the case where the FTIR is provided with the above-described HPLC, as shown in FIG. 39, a passage 87 connected with a detector 85 of a liquid chromatograph 86 comprising an eluted liquid tank 81, a liquid-sending pump 82, a sample-injecting portion 83, a partition column 84, detector 85 and the like is provided with a branching T-type joint 88 at an end thereof, tubes 89, 90 having suitable inside diameters being connected with joint 88, and one tube 89 being connected with an analyzer 91 while the other tube 90 is connected with an exhaust passage (not shown). In addition, reference numeral 92 designates a syringe for injecting the sample and reference numeral 93 designates a recorder.
In order to set a flow rate of a liquid flowing through each tube 89, 90, respectively, for example the following measures have been used:
1 The inside diameters of the tubes 89, 90 are changed;
2 Lengths of the tubes 89, 90 are changed; and
3 The tube 89 connected with analyzer 91 is provided with flow rate-setting means such as a needle valve.
However, according to the above described 1, the ratio of the flow rate of the liquid flowing through tube 89 to that of the liquid flowing through tube 90 is equal to the ratio of the square of the diameters but cannot be continuously changed.
According to 2, a continuous ratio can be obtained but not a length from detector 85 of liquid chromatograph 86 to analyzer 91 is changed and thus the time required for the movement of liquid is varied, but also long tubes are required in order to obtain an increased ratio.
In addition, according to 3, although not only a continuous ratio can be obtained but also the time required for the movement of liquid is not varied. For example, in case of an eluted liquid, a problem has arisen in that the current of the eluted liquid is made turbulent by the needle valve to expand a chromatogram, whereby it becomes difficult to maintain resolution power.